Development of genetic recombination technologies has provided drugs including a variety of proteins as an active ingredient. In particular, numerous drugs including antibodies as an active ingredient have been recently developed and commercialized. In addition, efficient production of these proteins in large-scale has become a more important issue in biopharmaceutical industry.
Generally, such proteins are produced by culturing recombinant cells in which a vector including a gene encoding a protein of interest is inserted. The culture broth includes impurities such as a wide variety of medium-derived components, host cell-derived components, protein-derived by-products or the like, in addition to the protein of interest. Thus, it is a very difficult and challenging task to achieve both the purification of the protein of interest by removing impurities to meet purity requirements for protein drugs as well as the efficient production of the protein of interest in large-scale.
In general, the protein purification method is carried out by a combination of different modes of chromatography. Chromatography is to separate the protein of interest from impurities, for example, based on charge, hydrophilicity, molecular size or the like.
In particular, when the protein of interest is an antibody, Protein A affinity chromatography or Protein G affinity chromatography is used as one of chromatography for purifying the antibody, by using binding property of Protein A or Protein G to the specific region of antibody such as Fc chain (Patent Document 1).
However, Protein A affinity supports generally used are very expensive in comparison to ion exchange supports or hydrophobic supports, and a vast amount of supports are needed for large-scale purification of antibodies in industrial drug productions or the like, resulting in an inevitable increase in the production costs.
Further, protein A affinity chromatography or protein G affinity chromatography is generally carried out in an adsorption mode of specifically adsorbing the antibody of interest onto the support, washing the adsorbing support to separate impurities, and finally eluting the antibody of interest from the support. In this regard, buffers used in the washing and eluting steps are different from each other, scale-up of chromatography apparatus brings out enlargement or complexity of the accompanying production facilities such as buffer tank, and moreover, manipulations become complicated. All of these factors are the cause of increasing production costs.
For these reasons, the drugs including proteins as an active ingredient require much higher production costs than drugs including small-molecule compounds as an active ingredient, which is a challenging problem. In other words, a reduction in the protein purification cost is demanded in this field.
On the other hand, it is known that enzymes secreted from host cells are included in a culture broth containing the protein of interest, and the protein of interest is degraded, modified, oxidized, or reduced by these enzymes during the protein purification process. For this reason, addition of enzyme inhibitors during the protein purification has been considered to prevent degradation, modification, oxidation, or reduction of the protein of interest (Patent Document 2). However, when the enzyme inhibitors are used during the protein purification, an additional process of removing the inhibitors is required, and moreover, certain inhibitors may affect the quality of the purified protein. Therefore, it cannot be said that the addition of inhibitors is the best way. Removal of host cell-derived enzymes is considered as one of the drastic methods to solve the problems, but it is essential to use chromatography. There is no simple method of removing these enzymes.
Activated carbon is an inexpensive natural material having extensive non-specific adsorption properties, and used as an adsorbent or as a decolorant in the industrial fields, such as the production of chemicals and foods, sewage or waste water treatment, water filtration, and production of small-molecule drugs. However, due to the extensive non-specific adsorption properties, it has been thought that there are difficulties in the use of activated carbon for high performance protein purification such as separation of the above-mentioned impurities. Thus, a method of purifying a protein using the activated carbon has not been known yet.